Variation in the Relative Isomer Abundance of Synthetic and Biologically Derived Phosphatidylethanols and Its Consequences for Reliable Quantification

Abstract Phosphatidylethanol (PEth) in human blood samples is a marker for alcohol usage. Typically, PEth is detected by reversed-phase liquid chromatography coupled with negative ion tandem mass spectrometry, investigating the fatty acyl anions released from the precursor ion upon collision-induced dissociation (CID). It has been established that in other classes of asymmetric glycerophospholipids, the unimolecular fragmentation upon CID is biased depending on the relative position (known as sn-position) of each fatty acyl chain on the glycerol backbone. As such, the use of product ions in selected-reaction-monitoring (SRM) transitions could be prone to variability if more than one regioisomer is present in either the reference materials or the sample. Here, we have investigated the regioisomeric purity of three reference materials supplied by different vendors, labeled as PEth 16:0/18:1. Using CID coupled with ozone-induced dissociation, the regioisomeric purity (% 16:0 at sn-1) was determined to be 76, 80 and 99%. The parallel investigation of the negative ion CID mass spectra of standards revealed differences in product ion ratios for both fatty acyl chain product ions and ketene neutral loss product ions. Furthermore, investigation of the product ion abundances in CID spectra of PEth within authentic blood samples appears to indicate a limited natural variation in isomer populations between samples, with the cannonical, PEth 16:0/18:1 (16:0 at sn-1) predominant in all cases. Different reference material isomer distributions led to variation in fully automated quantification of PEth in 56 authentic dried blood spot (DBS) samples when a single quantifier ion was used. Our results suggest caution in ensuring that the regioisomeric compositions of reference materials are well-matched with those of the authentic blood samples.

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Characterization of the molecular species of glycerophospholipids from rabbit kidney: An alternative approach to the determination of the fatty acyl chain position by negative ion fast atom bombardment combined with mass-analysed ion kinetic energy analysis

Biological Mass Spectrometry ◽

10.1002/bms.1200211207 ◽

1992 ◽

Vol 21 (12) ◽

pp. 655-666 ◽

Cited By ~ 11

Author(s):

Su Chen ◽

Ornella Curcuruto ◽

Silvia Catinella ◽

Pietro Traldi ◽

Gianpaolo Menon

Keyword(s):

Fast Atom Bombardment ◽

Acyl Chain ◽

Fatty Acyl ◽

Negative Ion ◽

Rabbit Kidney ◽

Fatty Acyl Chain ◽

Alternative Approach ◽

Ion Kinetic Energy

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Fatty acyl chain structure, orientational order, and the lipid phase transition in Acholeplasma laidlawii B membranes. A review of recent 19F nuclear magnetic resonance studies

Canadian Journal of Biochemistry and Cell Biology ◽

10.1139/o84-147 ◽

1984 ◽

Vol 62 (11) ◽

pp. 1134-1150 ◽

Cited By ~ 16

Author(s):

P. M. Macdonald ◽

B. D. Sykes ◽

R. N. McElhaney

Keyword(s):

Phase Transition ◽

Fatty Acids ◽

Nuclear Magnetic Resonance ◽

Acyl Chain ◽

Orientational Order ◽

Membrane Lipid ◽

Fatty Acyl ◽

Fatty Acyl Chain ◽

Lipid Phase ◽

Lipid Phase Transition

The orientational order parameters of monofluoropalmitic acids biosynthetically incorporated into membranes of Acholeplasma laidlawii B in the presence of a large excess of a variety of structurally diverse fatty acids have been determined via 19F nuclear magnetic resonance (19F NMR) spectroscopy. It is demonstrated that these monofluoropalmitic acids are relatively nonperturbing membrane probes based upon physical (differential scanning calorimetry), biochemical (membrane lipid analysis), and biological (growth studies) criteria. 19F NMR is shown to convey the same qualitative and quantitative picture of membrane lipid order provided by 2H-NMR techniques and to be sensitive to the structural characteristics of the membrane fatty acyl chains, as well as to the lipid phase transition. Representatives of each naturally occurring class of fatty acyl chain structures, including straight-chain saturated, methyl-branched, monounsaturated, and alicyclic-ring-substituted fatty acids, were studied and the 19F-NMR order parameters were correlated with the lipid phase transitions (determined calorimetrically). The lipid phase transition was the prime determinant of overall orientational order regardless of fatty acid structure. Effects upon orientational order attributable to specific structural substituents were discernible, but were secondary to the effects of the lipid phase transition. In the gel state, relative overall order was directly proportional to the temperature of the particular lipid phase transition. Not only the overall order, but also the order profile across the membrane was sensitive to the presence of particular structural substituents. In particular, in the gel state specific fatty acyl structures demonstrated a characteristic disordering effect in the membrane order profile. These various observations can be merged to provide a unified picture of the manner in which fatty acyl chain chemistry modulates the physical state of membrane lipids.

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The Role of Ceramide Metabolism and Signaling in the Regulation of Mitophagy and Cancer Therapy

Cancers ◽

10.3390/cancers13102475 ◽

2021 ◽

Vol 13 (10) ◽

pp. 2475

Author(s):

Megan Sheridan ◽

Besim Ogretmen

Keyword(s):

Cancer Therapy ◽

Acyl Chain ◽

Fatty Acyl ◽

Fatty Acyl Chain ◽

Cancer Cell Proliferation ◽

Bioactive Lipids ◽

Cellular Functions ◽

Proliferation Response ◽

Ceramide Synthases

Sphingolipids are bioactive lipids responsible for regulating diverse cellular functions such as proliferation, migration, senescence, and death. These lipids are characterized by a long-chain sphingosine backbone amide-linked to a fatty acyl chain with variable length. The length of the fatty acyl chain is determined by specific ceramide synthases, and this fatty acyl length also determines the sphingolipid’s specialized functions within the cell. One function in particular, the regulation of the selective autophagy of mitochondria, or mitophagy, is closely regulated by ceramide, a key regulatory sphingolipid. Mitophagy alterations have important implications for cancer cell proliferation, response to chemotherapeutics, and mitophagy-mediated cell death. This review will focus on the alterations of ceramide synthases in cancer and sphingolipid regulation of lethal mitophagy, concerning cancer therapy.

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Effect of fatty acyl chain length and structure on the lamellar gel to liquid-crystalline and lamellar to reversed hexagonal phase transitions of aqueous phosphatidylethanolamine dispersions

Biochemistry ◽

10.1021/bi00428a020 ◽

1989 ◽

Vol 28 (2) ◽

pp. 541-548 ◽

Cited By ~ 87

Author(s):

Ruthven N. A. H. Lewis ◽

David A. Mannock ◽

Ronald N. McElhaney ◽

David C. Turner ◽

Sol M. Gruner

Keyword(s):

Phase Transitions ◽

Chain Length ◽

Liquid Crystalline ◽

Hexagonal Phase ◽

Acyl Chain ◽

Fatty Acyl ◽

Fatty Acyl Chain ◽

Acyl Chain Length

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Effect of Acylglycerol Composition and Fatty Acyl Chain Length on Lipid Digestion in pH-Stat Digestion Model and SimulatedIn VitroDigestion Model

Journal of Food Science ◽

10.1111/1750-3841.13196 ◽

2015 ◽

Vol 81 (2) ◽

pp. C317-C323

Author(s):

Jin F. Qi ◽

Cai H. Jia ◽

Jung A. Shin ◽

Jeong M. Woo ◽

Xiang Y. Wang ◽

...

Keyword(s):

Chain Length ◽

Acyl Chain ◽

Fatty Acyl ◽

Fatty Acyl Chain ◽

Lipid Digestion ◽

Acyl Chain Length ◽

Ph Stat

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Phosphoglycolipid Profiling of Bacterial Endotoxins

Hungarian Journal of Industry and Chemistry ◽

10.1515/hjic-2018-0003 ◽

2018 ◽

Vol 46 (1) ◽

pp. 7-11 ◽

Cited By ~ 1

Author(s):

Anikό Kilár ◽

Ágnes Dornyel ◽

Viktor Sándor ◽

Ferenc Kilár ◽

Béla Kocsis

Keyword(s):

Lipid A ◽

De Novo ◽

Clinical Symptoms ◽

Phosphorylation Site ◽

Reversed Phase ◽

Fatty Acyl ◽

Structural Elucidation ◽

Bacterial Endotoxins ◽

Product Ions ◽

Tandem Mass Spectrometric

Abstract Much interest is at present focused on bacterial endotoxins, also known as lipopolysaccharides (LPS), as they are responsible for the development of clinical symptoms of Gram-negative sepsis which is the leading cause of death in intensive care units. Endotoxicity is associated with the special phosphoglycolipid part of LPS, termed lipid A. Main challenges in the structural elucidation of lipid A arise from its amphiphilic character and inherent heterogeneity. A mass spectrometrybased de novo method combined with reversed-phase liquid chromatography for the detailed structural characterization of complex lipid A mixtures (obtained by mild acid hydrolysis of LPS) from different bacterial sources has been developed. Tandem mass spectrometric measurements were performed with an electrospray-ionisation quadrupole time-of-flight (ESI-Q-TOF) mass spectrometer in both negative- and positive-ionization modes in order to explore fragmentation pathways. It was found that characteristic product ions in the positive-ion mode could be used for the unambiguous assignment of the phosphorylation site, whereas the use of both ionization modes provided consistent and/or complementary information about the fatty acyl distribution between the two glucosamine moieties of lipid A. Since the immunostimulatory (advantageous) vs. proinflammatory (endotoxic) effect of the lipid A is closely related to the fine chemical structure, our relatively simple structural elucidation strategy could offer great potential in the bioanalysis of native lipid A samples and lipid A-based vaccines

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Acyltransferase-mediated selection of the length of the fatty acyl chain and of the acylation site governs activation of bacterial RTX toxins

Journal of Biological Chemistry ◽

10.1074/jbc.ra120.014122 ◽

2020 ◽

Vol 295 (28) ◽

pp. 9268-9280 ◽

Cited By ~ 3

Author(s):

Adriana Osickova ◽

Humaira Khaliq ◽

Jiri Masin ◽

David Jurnecka ◽

Anna Sukova ◽

...

Keyword(s):

Acyl Chain ◽

Fatty Acyl ◽

Biologically Active ◽

Fatty Acyl Chain ◽

Acyl Carrier Proteins ◽

E Coli ◽

Rtx Toxins ◽

Wide Range ◽

Lysine Residues ◽

Acyl Chains

In a wide range of organisms, from bacteria to humans, numerous proteins have to be posttranslationally acylated to become biologically active. Bacterial repeats in toxin (RTX) cytolysins form a prominent group of proteins that are synthesized as inactive protoxins and undergo posttranslational acylation on ε-amino groups of two internal conserved lysine residues by co-expressed toxin-activating acyltransferases. Here, we investigated how the chemical nature, position, and number of bound acyl chains govern the activities of Bordetella pertussis adenylate cyclase toxin (CyaA), Escherichia coli α-hemolysin (HlyA), and Kingella kingae cytotoxin (RtxA). We found that the three protoxins are acylated in the same E. coli cell background by each of the CyaC, HlyC, and RtxC acyltransferases. We also noted that the acyltransferase selects from the bacterial pool of acyl–acyl carrier proteins (ACPs) an acyl chain of a specific length for covalent linkage to the protoxin. The acyltransferase also selects whether both or only one of two conserved lysine residues of the protoxin will be posttranslationally acylated. Functional assays revealed that RtxA has to be modified by 14-carbon fatty acyl chains to be biologically active, that HlyA remains active also when modified by 16-carbon acyl chains, and that CyaA is activated exclusively by 16-carbon acyl chains. These results suggest that the RTX toxin molecules are structurally adapted to the length of the acyl chains used for modification of their acylated lysine residue in the second, more conserved acylation site.

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Ectopic expression of ceramide synthase 2 in neurons suppresses neurodegeneration induced by ceramide synthase 1 deficiency

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1522071113 ◽

2016 ◽

Vol 113 (21) ◽

pp. 5928-5933 ◽

Cited By ~ 24

Author(s):

Stefka D. Spassieva ◽

Xiaojie Ji ◽

Ye Liu ◽

Kenneth Gable ◽

Jacek Bielawski ◽

...

Keyword(s):

Acyl Chain ◽

Strong Association ◽

Ectopic Expression ◽

Fatty Acyl ◽

Chemical Diversity ◽

Fatty Acyl Chain ◽

Length Variation ◽

Ceramide Synthase ◽

Hydrophobic Moiety

Sphingolipids exhibit extreme functional and chemical diversity that is in part determined by their hydrophobic moiety, ceramide. In mammals, the fatty acyl chain length variation of ceramides is determined by six (dihydro)ceramide synthase (CerS) isoforms. Previously, we and others showed that mutations in the major neuron-specific CerS1, which synthesizes 18-carbon fatty acyl (C18) ceramide, cause elevation of long-chain base (LCB) substrates and decrease in C18 ceramide and derivatives in the brain, leading to neurodegeneration in mice and myoclonus epilepsy with dementia in humans. Whether LCB elevation or C18 ceramide reduction leads to neurodegeneration is unclear. Here, we ectopically expressed CerS2, a nonneuronal CerS producing C22–C24 ceramides, in neurons of Cers1-deficient mice. Surprisingly, the Cers1 mutant pathology was almost completely suppressed. Because CerS2 cannot replenish C18 ceramide, the rescue is likely a result of LCB reduction. Consistent with this hypothesis, we found that only LCBs, the substrates common for all of the CerS isoforms, but not ceramides and complex sphingolipids, were restored to the wild-type levels in the Cers2-rescued Cers1 mutant mouse brains. Furthermore, LCBs induced neurite fragmentation in cultured neurons at concentrations corresponding to the elevated levels in the CerS1-deficient brain. The strong association of LCB levels with neuronal survival both in vivo and in vitro suggests high-level accumulation of LCBs is a possible underlying cause of the CerS1 deficiency-induced neuronal death.

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